HIV transmission within the body is primarily thought to occur via the direct interaction of virus positive-cells with uninfected target cells. Virus-positive cells are comprised of either cells that were previously infected or cells that have captured and maintained HIV in an infectious form on their cell surface. Impairing the cell-to-cell transmission of HIV is essential to stopping spread within the body and from person-to-person. We have constructed immunologically relevant model systems in cell culture to examine the cell-virus and cell-cell interactions that are required for virus transmission. Blood dendritic cells (DCs) efficiently capture and transmit HIV to CD4+ T lymphocytes, the main reservoir of virus infection in the body. DCs serve as immune system sentinels and patrol mucosal surfaces for invading pathogens to provide early warning to the adaptive immune system. Mucosal tissue exposure to HIV is the most frequent cause of HIV transmission worldwide. Because DCs are concentrated at these tissues, it is hypothesized that they are co-opted by HIV during mucosal transmission to gain access to the lymphatic system and seed infection of the CD4+ T cell compartment. DC-SIGN is a high-affinity HIV Env receptor expressed by activated B cells, a subset of blood macrophages, mucosal DCs, and in vitro differentiated monocyte-derived DCs (MDDCs). MDDCs and activated B cells treated with antibodies to DC-SIGN bind less HIV and are less effective in stimulating infection of CD4+ T cells. In addition, ectopic expression of DC-SIGN in transformed B cells enables efficient HIV capture and transmission to susceptible target cells. Cell biological analysis has revealed the concentration of HIV at synaptic junctions after DC contact with CD4+ target cells. It has been postulated that examination of DC-SIGNmediated HIV transmission by transformed cell lines could aid in the characterization of these processes. Transformed cells would allow more extensive biochemical characterization and genetic manipulation of the cellular sorting machinery, cell signaling processes, and cell-cell adhesion molecules required for synaptic transfer of HIV. Our group was the first to show the feasibility of modeling DC-SIGN-mediated transmission using transformed B cells in contrast to other cell types. We have used these differences to investigate cellular requirements for the DC-SIGN-mediated HIV transmission. These studies reveal that HIV traffics differently in cells that are permissive and restrictive to transmission and suggests that other cell factors regulate this process. In the course of these studies, we have found that GFP-labeled virus-like particles (VLPs) can be used to monitor the requirements of DC-SIGN- or DC-mediated transmission of HIV. We are now using VLPs to investigate residues of DC-SIGN required for interaction with HIV Env and to determine whether cell specific modification of DC-SIGN also regulates these interactions. These studies will increase our understanding of the cell-to-cell transmission of HIV, and they could provide the basis for rational design of antiviral drugs that prevent HIV use of DC-SIGN in virus transmission. [Corresponds to Project 1A in Dr. KewalRamanis May 2005 site visit report]